Local Expression of Indoleamine 2,3 Dioxygenase in Syngeneic Fibroblasts Significantly Prolongs Survival of an Engineered Three-Dimensional Islet Allograft

OBJECTIVE

The requirement of systemic immunosuppression after islet transplantation is of significant concern and a major drawback to clinical islet transplantation. Here, we introduce a novel composite three-dimensional islet graft equipped with a local immunosuppressive system that prevents islet allograft rejection without systemic antirejection agents. In this composite graft, expression of indoleamine 2,3 dioxygenase (IDO), a tryptophan-degrading enzyme, in syngeneic fibroblasts provides a low-tryptophan microenvironment within which T-cells cannot proliferate and infiltrate islets.

RESEARCH DESIGN AND METHODS

Composite three-dimensional islet grafts were engineered by embedding allogeneic mouse islets and adenoviral-transduced IDO–expressing syngeneic fibroblasts within collagen gel matrix. These grafts were then transplanted into renal subcapsular space of streptozotocin diabetic immunocompetent mice. The viability, function, and criteria for graft take were then determined in the graft recipient mice.

RESULTS

IDO-expressing grafts survived significantly longer than controls (41.2 ± 1.64 vs. 12.9 ± 0.73 days; P < 0.001) without administration of systemic immunesuppressive agents. Local expression of IDO suppressed effector T-cells at the graft site, induced a Th2 immune response shift, generated an anti-inflammatory cytokine profile, delayed alloantibody production, and increased number of regulatory T-cells in draining lymph nodes, which resulted in antigen-specific impairment of T-cell priming.

CONCLUSIONS

Local IDO expression prevents cellular and humoral alloimmune responses against islets and significantly prolongs islet allograft survival without systemic antirejection treatments. This promising finding proves the potent local immunosuppressive activity of IDO in islet allografts and sets the stage for development of a long-lasting nonrejectable islet allograft using stable IDO induction in bystander fibroblasts.Endocrine replacement therapy by islet transplantation represents a feasible and attractive alternative therapeutic approach for treating type 1 diabetes (1,2). Despite improvement of allogeneic islet engraftment using systemic immunosuppression, islet transplantation is still limited by high rates of rejection. Furthermore, some immunosuppressive agents are prodiabetogenic and associated with adverse side effects (3–6). Finding more efficient and less harmful strategies to protect islet graft is therefore required for improving islet transplantation outcome.Localized expression of immunoregulatory factors using gene transfer to graft is a feasible method to provide an immunoprivileged microenvironment and consequently improves graft survival. Such an on-site delivery system results in more potent local immunosuppression with less systemic side effects (7–9).IDO is a cytosolic enzyme that catalyzes essential amino acid l-tryptophan to kynurenine (10) and has profound effects on T-cell proliferation, differentiation, effector functions, and viability (11). Both the reduction in local tryptophan concentration and the production of immunomodulatory tryptophan metabolites contribute to immunosuppressive effects of IDO (12,13). Broad evidence implicates IDO and the tryptophan catabolic pathway in generation of immune tolerance to antigens in tissue microenvironments. In particular, the role of IDO in fetal tolerance in mammalian pregnancy (14,15), immunologic tolerance to tumors (16,17), and self-tolerance has been documented (18,19). The unique immunoregulatory function of IDO substantiates the application of this enzyme as a strategy to suppress alloimmune responses in transplantation.Our research group has shown that overexpression of IDO in fibroblasts suppresses immune response and improves outcome of skin grafts (20–25) and that bystander IDO-expressing fibroblasts suppress immune response to allogeneic mouse islets in vitro (26). Furthermore, in a recent study we showed that mouse islets and fibroblasts are selectively resistant to IDO-mediated activation of nutrient deficiency stress (27). Here, we engineered a three-dimensional composite islet allograft equipped with IDO-expressing fibroblasts and examined whether local expression of IDO, conferred by adenoviral-mediated gene transfer to bystander syngeniec fibroblasts, prevents the rejection of islet allograft. Our approach here is novel compared with other studies that examined the suppressive effect of IDO in islet transplantation (28,29) because 1) bystander syngeneic fibroblasts were used as the target of gene transfer instead of islets to avoid deleterious effects of adenovirus infection on islets (30–32), 2) islets were embedded within an extracellular matrix that by itself improves islet function and viability (33,34), and 3) cotransplanted fibroblasts are more than just a source of IDO and can enhance islet physiological competence (35,36).     

Correlation between testosterone, gonadotropins and prolactin and severity of negative symptoms in male patients with chronic schizophrenia

The aim of this study was to evaluate the relationship between plasma levels of testosterone, FSH, LH and prolactin and the severity of negative symptoms in patients with chronic schizophrenia. Fifty-four male inpatients with chronic schizophrenia participated in this cross-sectional study. Twenty-five age-matched men without a history of psychiatric disorders or endocrine illnesses were used as controls. All patients were on risperidone 4 mg/day or haloperidol 10 mg/day and anticholinegic medication, biperidine 3 mg/day. The patients were assigned to groups with predominant negative and nonpredominant negative symptoms on the basis of their scores on the Positive and Negative Syndrome Scale (PANSS). Plasma levels of testosterone and free testosterone in the patients with predominant and nonpredominant negative symptoms were significantly lower than those in normal controls. Furthermore, plasma levels of FSH and LH, in the patients with predominant negative symptoms but not in the nonpredominant negative symptoms, were significantly lower than those in the normal controls. In contrast, plasma level of prolactin in the predominant negative symptoms group but not in the nonpredominant negative symptoms group was significantly higher than the aged matched normal males. Significant inverse correlation between negative subscale scores of PANSS and plasma levels of testosterone and free testosterone in the patients with predominant negative symptoms were detected. There was also a positive correlation between prolactin plasma levels and negative subscale scores. The present study indicates that assessment of sex hormones and function of hypothalamic-pituitary-gonadotropin axis could be an important biological marker for the severity of negative symptoms in schizophrenia and these findings may change the present pharmacotherapy for negative symptoms based on neuroendocrinology profiles of patients with schizophrenia.     

Is the role of Chlamydia trachomatis underestimated in patients with suspected reactive arthritis?

Reactive arthritis is usually caused by bacteria of either the enteric or genital tracts. In the genital tract, Chlamydia trachomatis is perhaps the only aetiological agent. In Iran, newer evidence suggests that as C. trachomatis is more commonly found in the general population than was previously believed, its role in reactive arthritis may well be currently overlooked. In this review, as well as emphasizing the potential role of C. trachomatis in reactive arthritis in patients from developing countries, we also make recommendations for further clinical studies to determine its prevalence. Moreover, we also stress the need for standardization of new testing methodologies for C. trachomatis, including the use of new commercial systems in an attempt to determine a truer picture of chlamydial infection in reactive arthritis.     

Tumoral cell mtDNA approximately 8.9 kb deletion is more common than other deletions in gastric cancer

BACKGROUND: The aim of the study was to clarify the role of deletion of mitochondrial DNA (mtDNA) in gastric carcinogenesis and to determine prevalence of mitochondrial deletions in different regions of tumoral tissue in comparison with adjacent non-tumoral tissue in gastric cancer. METHODS: In order to investigate whether a high incidence of mutations exists in mtDNA of gastric cancer tissues, we screened five regions of the mitochondrial genome by PCR amplification, Southern blot and DNA sequence analysis. RESULTS: Of 71 cancer patients, the approximately 8.9 kb deletion was detected among different deletions in 9 cases (12.67%) of the tumoral tissues and 1 case (1.40%) in non-tumoral tissues that were adjacent to the tumors. Level of the 8.9 kb deletion has been found to be more than other deletions in tumoral tissues. CONCLUSIONS: The approximately 8.9 kb deletion has an obvious correlation with age and histological type. These data suggest that the approximately 8.9 kb deletion in mtDNA may play an important role in gastric carcinogenesis.     

The protean manifestations of varicella-zoster virus vasculopathy

Varicella-zoster virus (VZV) vasculopathy in the central nervous system (CNS) affects large and small cerebral vessels. Large-vessel disease is most common in immunocompetent individuals, whereas small-vessel disease usually develops in immunocompromised patients. In some patients, both large and small vessels are involved. Neurological features are protean. Neurological disease often occurs months after zoster and sometimes without any history of zoster rash. Magnetic resonance imaging (MRI) scanning, cerebral angiography, and examination of cerebrospinal fluid (CSF) with virological analysis are needed to confirm the diagnosis. VZV vasculopathy patients do not always have VZV DNA in CSF, but diagnosis can be confirmed by finding anti-VZV antibody in CSF, along with reduced serum/CSF ratios of VZV immunoglobulin G (IgG) compared to albumin or total IgG. When VZV vasculopathy develops months after zoster, antiviral treatment is often effective.     

Varicella in Americans from NHANES III: implications for control through routine immunization

At the time of varicella vaccine introduction in the United States, an estimated 4 million episodes of varicella occurred annually. This survey of varicella seroprevalence is the first to describe immunity to a vaccine-preventable disease prior to vaccine introduction in the United States population. The objective of this analysis is to describe patterns of naturally-acquired varicella and understand characteristics associated with infection in the varicella vaccine-naive United States population. A nationally representative cross-sectional health examination survey that included venipuncture was conducted among 21,288 U.S. participants aged 6 years and older from 1988 through 1994. Serologic evidence of varicella-zoster virus infection was measured by enzyme immunoassay of varicella-zoster virus-specific IgG antibody. The seroprevalence of IgG antibody to varicella-zoster virus increased from 86.0% in children aged 6 through 11 years to 99.6% in adults aged 40 through 49 years. Immunity to varicella remained at 99% or higher in Americans aged 50 years and older. Among persons aged 6 through 19 years, non-Hispanic black children were 40% less likely to be seropositive compared with white children (odds ratio [OR], 0.6; 95% confidence interval [CI], 0.4-0.8). Among young adults aged 20 through 39 years, women with a history of live birth (OR, 4.3; 95% CI, 2.1-8.7) and married men (OR, 2.7; 95% CI, 1.2-5.7) were more likely to have naturally-acquired immunity to varicella. This study found that, prior to use of varicella vaccine in the United States, age, race, and marital characteristics were independently associated with naturally acquired varicella. Future varicella serosurveys in Americans will provide essential information to interpret the population impact of varicella vaccine.     

Clinico-pathological features of thyroid cancer as observed in five referral hospitals in Iran--a review of 1177 cases

In this study some of the clinical and pathological aspects of thyroid carcinomas in Iran are defined. Until fairly recently, Iran was an area of endemic iodine deficiency. The medical records of 1  177 confirmed cases of thyroid cancer treated over a 15-year period at 5 tertiary referral centers were reviewed. The female/male ratio was 1.8/1. Mean age was 42.8 ±0.9 years, male patients being significantly older. The most common clinical presentation (initial manifestation) was a central neck mass; 28.6% of the tumors had metastasized, usually to the cervical lymph nodes, by the time the patient presented with disease. The mean size of non-metastatic tumors was 5.0 cm at their longest diameter. The distribution of tumors in our study did not reflect the expectations for an iodine-deficient area, where follicular thyroid carcinoma is common, but rather what is seen in iodine-rich areas. Papillary and follicular types accounted for79.7% and 8.8% of cases, respectively.     

Earliest time of change in QT dispersion after stenting in patients with single vessel coronary artery disease

Dispersion of the QT interval (QTd) is a measure of inhomogeneity of ventricular repolarization, and its prolongation may provide a suitable substrate for life-threatening ventricular arrhythmias. The present study was performed to determine the onset time of change in the corrected QT (QTc) interval and QTd in patients with stable angina and single vessel coronary artery disease. Electrocardiograms of 60 patients with successful stenting, obtained 1 h before and 1 h, 6 h, 12 h and 24 h after the procedure were analyzed. The QTc interval, QTc maximum, QTc minimum and QTd were measured. All electrocardiograms were scanned, and then underwent computer-based analysis. There was a significant reduction in the mean QTc interval as early as 12 h after the procedure (from 474±41 ms to 460±31 ms; P<0.001), which persisted to the 24 h follow-up. This was associated with a significant reduction in mean QT maximum (from 496±31 ms to 418±66 ms; P<0.001) and a significant prolongation in mean QT minimum (from 403±21 ms to 444±12 ms; P<0.001) at the same time intervals. Therefore, successful stenting of coronary arteries in patients with single vessel coronary artery disease and stable angina decreases QTd as early as 12 h after the procedure. This phenomenon may be the result of improved regional myocardial circulation, and reduced ischemia. A persistently low QTd in the following months may therefore have prognostic significance, and can be used as a noninvasive marker of stent patency. Further studies are necessary to define the clinical applicability of QTd in the assessment of long-term stent patency in such patients.     

Total antioxidant capacity and levels of epidermal growth factor and nitric oxide in blood and saliva of insulin-dependent diabetic patients

BACKGROUND: The aim of this study was to examine how type 1 diabetic patients have altered levels of lipid peroxidation, antioxidant defense, NO and EGF in their plasma and saliva. We tested the differences in lipid peroxidation level, antioxidant power, and concentrations of epidermal growth factor (EGF) and nitric oxide (NO) in saliva and blood of type 1 diabetic subjects in comparison to healthy control subjects. METHODS: Nineteen subjects with type 1 diabetes mellitus and 19 healthy age- and sex-matched control subjects were included in the study. Blood and saliva samples were obtained and analyzed for thiobarbituric reactive substances (TBARS) as a marker of lipid peroxidation, ferric reducing ability (total antioxidant power), EGF and NO levels. RESULTS: TBARS levels did not show a significant difference between the two groups. Analysis of antioxidant power revealed that saliva and plasma of diabetic patients had more antioxidant power (p <0.01) than the healthy control population (107 +/- 10.35 vs. 11.14 +/- 4.66 and 192 +/- 12.3 vs. 142 +/- 15.2 mmol/L, respectively). Concentration of EGF was increased (p <0.01) in saliva whereas it was reduced (p <0.01) in plasma of diabetic patients in comparison to those of healthy subjects (2423 +/- 322 vs. 1513 +/- 341 and 125 +/- 14 vs. 346 +/- 60 pg/mL, respectively). NO level increased in both saliva and plasma of diabetic patients in comparison to those of healthy subjects (46.61 +/- 7 vs. 72.89 +/- 13 and 62.11 +/- 4.6 vs. 76.25 +/- 5 micromol/L, respectively). Blood HbA1c (%) of patients was significantly higher than that of controls (8.3 +/- 0.32 vs. 5.4 +/- 0.24, p <0.01). CONCLUSIONS: Existence of increased total antioxidant power in the presence of normal lipid peroxidation in plasma and saliva of type 1 diabetic patients indicates the existence of oxidative stress. Increased salivary EGF and NO levels in association with elevated TAOP is interesting and should be further studied.